The overall goal of this proposal is to develop a novel combination therapeutic strategy for treatment of pancreatic cancer using our newly developed anti-HER3 antibodies, which target both ligand-dependent and -independent pathways of HER3 activation. The central hypotheses are: (1) the extracellular domain of HER3 has at least two critical functional domains responsible for activation of HER3: one is responsible for the ligand binding and mediates HER3 activation in a ligand-dependent fashion, and another mediates and maintains HER3 activation in a ligand-independent fashion; (2) the activation of HER3 via both ligand-dependent and independent pathways are equally important for the development of resistance of pancreatic cells to EGFR and/or HER2 targeted therapies; and (3) the blockade of both pathways is required for a complete inactivation of HER3, which further would greatly enhance the efficacy of other targeted therapies such as erlotinib and lapatinib. The hypotheses were raised and have been supported by identification and characterization of a novel monoclonal anti-HER3 antibody, clone: 1A5, which primarily inhibits phosphorylation of HER3 in a ligand-independent fashion. The importance of the ligand-independent activation of HER3 was demonstrated by in vitro and in vivo anti-tumor efficacy of 1A5 alone and in combination with erlotinib or lapatinib. Furthermore, both HER3 activation pathways exist in most HER3+ pancreatic cancer cell lines we have tested, which appear to be associated with HER2 activation, and the resistance to erlotinib and lapatinib. As several anti-HER3 antibodies targeting ligand-dependent activation are under early phase clinical evaluation, our preliminary data raise a concern that only inhibition of the ligand-dependent activation of HER3 is not sufficient to override its agonistic activity to other receptors in the ERBB family, particularly, EGFR and HER2. The feasibility of this study is supported by the availability of both anti-HER3 antibodies and a novel assay system for quantitative measurement of the expression and function of the three major ERBB receptors: EGFR, HER2 and HER3, which allows us to determine the biological functions of interaction among the three receptors and the molecular mechanisms of the proposed combination therapy. Two Specific Aims will be addressed: AIM 1 will determine the role of the ligand-independent activation of HER3 in the activation of EGFR and HER2 in a panel of human pancreatic cancer cell lines and fresh pancreatic cancer tissues; and AIM 2 will determine the therapeutic efficacy of anti-HER3 in combination with erlotinib/lapatinib and gemcitabine.